Preservation, fixation, and staining: Delay in specimen preparation and fixation can result in poor preservation and suboptimal cytologic detail (Fig. 11.1a, b). This is particularly true of cerebrospinal fluid (CSF) specimens, in which cells rapidly degenerate at room temperature, making interpretation difficult or impossible. Cytomorphology is also affected by differences in fixation and staining techniques. As illustrated in Fig. 11.1c, d, the nuclear and cytologic features of the cells in the alcohol-fixed Giemsa-stained preparation differ from those of the cells in the air-dried Giemsa-stained preparation from the same specimen. Moreover, an examiner accustomed to one method may view differences related to preparation by another method as artifacts [2]. Table 11.2 compares cytomorphologic features of fresh alcohol-fixed and air-dried preparations.

Thick preparations: Thick smears or cytospins with excess material can result in poorly fixed and poorly stained areas that are difficult or impossible to evaluate morphologically. Moreover, even in areas with adequate fixation and staining, cytologic features may be partly or completely obscured (Fig. 11.2).

Cytospin preparation artifact : The centrifugal force used in cytospin preparations can result in cellular distortion, including nuclear irregularity, cellular molding (Fig. 11.3a, b), and edge artifact (Fig. 11.3c).

Degenerative changes: In some specimens, particularly urine, degenerative changes can result in nuclei with glassy or smudged chromatin that mimic viral inclusions. Similarly, mononuclear inflammatory cells with degenerative changes can mimic small round blue cell tumors, and degenerating benign epithelial and mesenchymal cells can be misinterpreted as malignant due to dark, condensed chromatin, nuclear irregularities, and increased nuclear to cytoplasmic ratios. Loss of chromatin detail, breaks in the nuclear membrane, and vacuolated or disrupted cytoplasm provide important clues for distinguishing degenerative changes from viral infection and/or malignancy.

Air bubbles and water bubbles: Air bubbles can result from hasty coverslipping or areas of thick, unevenly distributed cytologic material. Water bubbles can be a result of mounting a wet slide (Fig. 11.3d).

Dilution with blood and/or contamination with bone marrow: Excessive dilution with blood reduces the number of lesional cells and may delay fixation. Bone marrow or extramedullary hematopoietic cells in CSF and other specimens can create diagnostic difficulties, as early myeloid and/or erythroid precursors may be misconstrued as malignant cells (Fig. 11.4a, b). Usually megakaryocytes are not seen in CSFs with bone marrow contamination; however, in specimens with extramedullary hematopoietic cells, megakaryocytes are known mimickers of malignant cells, due to their large size and hyperchromatic, lobated nuclei.

Necrosis and crush artifact: “Naked nuclei” and distorted and crushed cells are features of cellular fragility, but can also be present in necrosis. Apoptotic and degenerative changes are often present in necrosis and tend to be more striking in specimens with delayed processing (Fig. 11.4c). Necrosis can also appear as a background of acellular amorphous or granular debris on the stained slide.

Thermal factors: Lasers or cold knives are used in some surgeries and produce characteristic artifacts in tissue that may confound interpretation of touch and squash/smear preparations from these specimens. Figure 11.4d illustrates freezing artifact in a case of mesenchymal chondrosarcoma. Fragmentation and microspherocytes of red blood cells can also be a clue to thermal artifact (Fig. 11.5).